A cell culture device is described comprising a gas permeable vessel which contains one or more self contained carbon dioxide generators and which is attached to a closure closing a gas permeable cell culture flask. Located in the closure is a gas permeable insert which defines a gas permeable opening through which the gases from the atmosphere of the vessel produced by the carbon dioxide generators can communicate with the gases in the atmosphere of the flask. This invention allows the rapid and uniform equilibration of the atmosphere of the flask with the controlled atmosphere of the vessel while still providing a closed system which prevents the entry of microbial organisms into the flask. This invention provides a controlled atmosphere for up to 24 hours that maintains the pH of the culture medium for optimal growth of cells growing in the culture medium in the flask. This invention allows for continuous microscopic observation of cells during this period and for long term exposure of cells to various physical or chemical agents.